In this merit extension proposal, we are requesting funds to extend the critical findings of SI P release and signaling in the vasculature. The first major aim ofthe research is to understand how SIP metabolic enzymes are regulated in the vascular endothelial cells to impact local (autocrine) as well as systemic SIP signaling. Secondly, we will fully explore the basic mechanisms and physiological impact of the SIP signaling system as a sensor of biomechanical shear stress on the endothelial cells. First, we hypothesize that Sphkl in the endothelial cell is primarily involved in local/autocrine signaling mode important for endothelial cell homeostatic and angiogenic reactions. We will derive an endothelial cell-specific inducible knockout of Sphkl and examine angiogenesis, endothelial cell patterning, pathologic angiogenesis, vascular permeability and inflammation. To examine the effect of systemic SI P, which is derived primarily by red blood cells, we will examine endothelial cell functions in the red blood cell-specific Sphkl knockout. Localized SI P signaling will be defined at the morphologic and ultrastructural level by immunolocalization studies in vivo as well as in primary cells. Second, we will explore the novel hypothesis that SIP metabolism and signaling constitutes a novel shear sensor/ signaling module in the endothelial cells. Mechanisms of how SIP is formed and released will be explored in detail in cultured vascular endothelial cells. In particular, we will focus on mechanisms that down regulate SIP lyase enzyme. Interestingly, laminar shear-induced downstream signaling mechanisms are critically dependent on SI PI receptor signaling, suggesting that this receptor is a sensor of physiological shear stress. We will explore this mechanism in detail using cultured endothelial cells. We anticipate that these experiments will allow us to better understand the role of sphingolipid metabolism and signaling in vascular endothelial cells. Since SIP receptor modulators have entered the therapeutic era, this knowledge is likely to have important clinical utility.